In the present study, the synthesized Multi-Walled Carbon Nanotubes (MWCNTs) were chemically treated with a mixture of acids to produce functionalized MWNTs. The functionalized MWCNTs were characterized by X-Ray Diffraction Analysis (XRD), Zeta potential and Field-Emission Scanning Electron Microscopy (FE-SEM). The X-ray diffraction reveals the MWCNTs average crystal size of the R-MWCNTs and F-MWCNTs were about (3.27 and 3.19) nm, respectively. FESEM images show the formation of R-MWCNTs that appears as cylindrical tubes and aggregated tubes with each other, while the F-MWCNTs appear as less aggregated and tangled clusters than R-MWCNTs. Zeta potential measurements showed that the measurement of the R-MWCNT shows a low negative value -9 mV and F-MWCNT, it was found that the zeta potential value is up to -29 mV. The antibacterial activity was studied against E. coli and P. aeruginosa bacteria, and indicated the highest growth inhibition zones (IZ) of F-MWCNTs as compared with R-MWCNTs against E. coli and Pseudomonas aeruginosa, respectively.